Mechanisms of Protective!Immunity after Dengue Natural Infections and Vaccination OVERALL P01 SUMMARY The four dengue virus serotypes (DENV1-4) cause the most important mosquito-borne viral disease of humans, with 100 million cases annually. The mechanisms by which the host immune response to DENV provides either protection against or enhancement of a subsequent infection with a different DENV serotype are not fully understood, and this has been a major concern in vaccine development and implementation. The poor results of the first registered dengue live attenuated vaccine highlight the critical need to better understand the immune response to natural DENV infections and vaccine candidates and to identify robust correlates of protection. Our current Program Project has been very successful, generating >50 high-level publications in 3.5 years and producing key findings that are timely, topical, and impactful. Here, we propose to identify immune mechanisms responsible for outcomes of DENV infection and vaccination, comprehensively exploring both the antigenic specificity and Fc functionality of antibodies as well as the quality of T cell responses in the context of long-term ongoing clinical and epidemiological studies of natural DENV infections and vaccines. We integrate classical cutting-edge molecular genetic and systems serology approaches to systematically examine both antigen specificity and Fc characteristics and effector functions of antibodies, as well as T cell attributes, that predict protection or pathogenesis. We propose a coordinated P01 Program that includes four projects: 1) Quality of B Cell and Antibody Responses to Natural Dengue Virus Infections; 2) Dengue Vaccines: Linking Vaccine-Induced Antibody Responses to Protective or Disease-Enhancing Immunity; 3) Quality of T Cell Responses Following Dengue Virus Natural Infections and Live-Attenuated Dengue Virus Vaccination, and 4) Genetic and Structural Basis for Human Antibody Inhibition of Dengue Viruses. The P01 is highly synergistic in that samples from the same individuals, as well as specific assays, reagents and methodologies, are shared among the Projects, which are supported by an Administrative Core, Computational Biology and Statistical Modeling Core, and Clinical and Data Management Core. We will leverage unique sample sets from the longest continuous cohort study of dengue, in Nicaragua, as well as a cohort study following recipients of the Dengvaxia vaccine in the Philippines and human challenge studies of the NIH live attenuated vaccine. We have have expanded our existing P01 Consortium of world-renowned investigators with extensive experience and on-going programs in dengue clinical, immunological, and virological research and vaccine development ? thus ensuring a continued high-quality successful research program. All the partners have a long-standing record of collaboration and numerous joint publications (>225). The Specific Aims are: 1) Define the antigenic repertoire and Fc characteristics of antibodies that predict outcome of subsequent DENV infection as well as the long-term humoral immune response; 2) Link vaccine-induced antibody responses to specific outcomes such as protection, vaccine failure and vaccine-primed severe disease; 3) Demonstrate that quality of T cell responses influences the clinical outcomes of infection and vaccination and modulates the quantity and quality of B cell responses; 4) Comprehensively define the protective antigenic landscape of DENV1-3 type-specific as well as DENV cross-reactive monoclonal and polyclonal antibodies. The overarching premise is that the accumulation of immunologic data from natural DENV infections, vaccine cohorts, and human challenge studies, coupled with state-of-the-art computational modeling and data integration, will enable identification of new immune correlates that predict protective immunity or immune enhancement, providing critical metrics for evaluating existing and second-generation DENV vaccines.